Inhaled Oxytocin and HPA Axis Reactivity

The Mood, Mother and Infant Study is a prospective observational cohort study that began
enrollment in May of 2013. Women were recruited in the 3rd trimester of pregnancy and
followed prospectively through 12 months postpartum. Mother-infant pairs who completed the
12-month visit will be invited to participate in the current Psychobiology of Resilience in
Maternal-Child Pairs follow-up study. The trial described here is a randomized controlled
trial embedded within the Psychobiology of Resilience study.

Non-pregnant women will be randomized to either 24 IU of nasal oxytocin (OT) or placebo. The
Investigational Drug Service (IDS) will use a random number generator to prepare a
randomization table. Participants will be block randomized by risk status at enrollment in
the Mood, Mother and Infant (MMI) study, as verified by structured clinical diagnostic
interview (No history of depression or anxiety, Past depression or anxiety, current
depression or anxiety). Both participants and study personnel will be blinded to allocation
group. At the end of the protocol, participants will be asked which condition they believed
they were in ('oxytocin,' 'control,' 'not sure') to ascertain success of blinding. Forty
minutes after treatment, women will undergo the Trier Social Stress Test (TSST), comprised of
a speech task and a math task; the TSST reliably induces large and consistent HPA and
cardiovascular responses. The TSST is administered as follows: Pre-Task Instructions: (5
minutes) Subjects will be introduced to 3 people (the 'selection committee') and asked to
assume the role of a job applicant. Anticipation Period: The subject prepares her speech for
3 minutes in the presence of the committee. Speech: The committee asks the subject to deliver
her talk for 5 minutes while being video and audio-recorded. If the subject finishes early,
the committee responds with prepared questions to ensure that she speaks for the entire 5
minutes. These questions are designed to be non-harassing but to create a feeling of lack of
predictability/controllability (e.g., "Do you have any enemies?") Serial Subtraction (PASST):
The committee will ask the subject to subtract the number 7 from 2000 as quickly and
accurately as possible for 5 minutes. For each mistake, the committee says "Stop -- mistake
-- start over at 2000." Stress Recovery: The subject sits quietly alone for 20 minutes.

Blood will be collected at baseline, during the speech and math tasks, and at 10 and 20
minutes of recovery, as HPA-axis responses are reliably found 10-30 minutes following the
TSST. Evidence regarding optimal timing of stress testing is conflicting. Visits will be
scheduled for 1 pm to increase likelihood of detecting a stress response unopposed by the
circadian influence, based on the experience of investigators in our laboratory and published
studies of postpartum women. These investigators have found menstrual cycle phase does not
affect TSST results; therefore, the date of last menstrual period and hormone use will be
recorded, but visits will not be scheduled based on cycle phase.

This study will follow-up the existing Mood, Mother and Infant (MMI) prospective
longitudinal cohort (R01HD073220), comprised of 222 mother-infant dyads who were recruited
between May 2013 and April 2017 and completed the 12-month MMI visit. In the MMI study, 222
mothers ages 18-45 and their infants were enrolled. Participants were recruited from
community clinics in the third trimester of pregnancy and continued to participate in the
study through 12 months postpartum. At the 12-month visit, mothers were invited to continue
to be followed via online surveys at 6-month intervals; more than 80% of women who have
completed the MMI study to date have continued to participate. Enrolled participants in the
MMI study met the following inclusion and exclusion criteria:

Inclusion Criteria:

1. Singleton pregnancy;

2. Intention to breastfeed (due to the centrality of breastfeeding to the oxytocin
assessment);

3. Intention to remain within 40 miles of the University of North Carolina - Chapel Hill
through infant's first birthday;

4. Ability to communicate in English.

Exclusion Criteria:

1. Maternal diagnosis of Axis I disorders other than unipolar depression or anxiety
disorders. Women with a history of bipolar disorder were excluded, given their
increased risk of postpartum psychosis.

2. Active substance abuse at enrollment in the 3rd trimester of pregnancy (Tobacco,
alcohol, illicits);

3. Major congenital anomaly;

4. Chronic medication/medical condition contraindicated for breastfeeding;

5. Current use of tricyclic antidepressants, which alter cortisol and heart rate
variability.

At enrollment, all participants underwent a Structured Clinical Interview Non-Patient
version (SCID-NP).

Inclusion Criteria for Inhaled Oxytocin and HPA Axis Reactivity, a substudy of the
Psychobiology of Resilience in Mother-Child Pairs follow-up study: 1) Participated in the
MMI study 2) Both mother and child willing and able to participate in the 6-year follow-up
visits 3) Not pregnant, verified by urine pregnancy test on day of study visit.